An important function of the vertebrate kidney is the excretion of potentially toxic chemicals such as waste products of cellular metabolism, xenobiotics and xenobiotic metabolites. This occurs primarily in the proximal tubule where specific renal transport systems remove organic anions and organic cations from peritubular capillaries and move them across the tubular epithelium into the lumen. We are using comparative renal models (proximal tubules from lower vertebrates and invertebrates and mammalian renal cells in culture) in combination with fluorescence microscopy (conventional and confocal), video imaging, intracellular microinjection and isolated membrane vesicle techniques to define the cellular mechanisms that underlie xenobiotic excretion. We have found that organic anions and cations are distributed over two compartments within kidney, liver and choroid plexus cells: one diffuse and cytoplasmic, and the second punctuate and vesicular. Sequestration in intracellular vesicles is not a result of endocytosis, but rather of active uptake from the cytoplasm. Moreover, in proximal tubule, organic anion-containing vesicles appear to move through the cytoplasm in the secretory direction on micro-tubules; this movement contributes substantially to the rate of net secretion. In addition, imaging studies have provided the first demonstration in intact renal tubules of xenobiotic (cyclosporin and daunomycin) secretion mediated by the multidrug resistance (MDR) transporter. This ATP-driven pump is the third major excretory transport system with broad specificity limits present in the renal proximal tubule. Future studies will focus on: 1) characterizing vesicular transport mechanisms driving xenobiotic secretion in proximal tubule, especially interactions with the cytoskeleton, 2) defining the role of the MDR transporter in xenobiotic secretion, 3) identifying the mechanisms that regulate xenobiotic secretion, and 4) characterizing analogous systems in other specialized epithelia that accumulate and transport organic anions and cations, e.g., liver and choroid plexus.